Further properties and the mechanism of action of homogeneous inositol 1-phosphate synthase (EC 5.5.1.4) from rat testis cytosol have been examined. Inosose-2 1-phosphate has been shown indirectly, but conclusively, to be the cyclic intermediate which is reduced presumably by NADH to the product. Antibody to the enzyme prepared in rabbits cross reacts with all rat tissue synthases studied. The immunoprecipitated enzyme is 60% as active as the soluble enzyme. Overall purification of the enzyme, described previously, has now been determined at 450-fold with introduction of the immunochemical assay.